The rapidly advancing technology of photon diffusion has afforded the opportunity to quantify hemoglobin saturation and blood content in the exercising limb, whilst previously only "trend indications" were possible. The quantitation of hemoglobin/myoglobin spectroscopy of the exercising limb is now possible with greatly simplified direct readout, time-simplification and increased precision of the phase modulation spectroscopy (PMS) enables the use of light emitting diodes, on the one hand, and PIN diode detectors on the other, to make this direct measurement of pathlength directly available for wide range of applications to exercising limbs. The 1H NMR determined myoglobin deoxygenation enables a quantification of its contribution to the optical readout of Hb+Mb and at the same time, the capillary bed-cytosol oxygen gradient in exercising tissue. The proposed activity is to be greatly extended with these new technologies as critically evaluated in Aim 1 by a sophisticated "muscle model" of oxygen delivery to tissue with appropriate scattering material and a model capillary bed for a) calibrating new technologies and b) evaluating their performance in imaging localized volumes in exercising skeletal muscle. Aim 2 is to compare mitochondrial hypoxia with deoxygenation states of hemoglobin and myoglobin in an animal muscle model in which the optical methods, 1H NMR, and freeze-trap redox scanning of the mitochondrial redox state are compared, the latter is proposed to be a "Gold Standard" of mitochondrial hypoxia. The gradient between the capillary to tissue and tissue to mitochondrial will be established and measured in a way that is transferable to human subjects through the use if 1H NMR both in Aims 2 and 3, and optical methods. Aim 3 proposes the application of the 1H NMR, and PMS/TRS methods to the evaluation of the desaturation of hemoglobin and myoglobin in exercising limbs of humans in a variety of protocols in individuals of exercise capabilities ranging from elite performers, normal subjects, to those with metabolic and genetic decreases of bioenergetic capability. Thus, the impact of exercise capability upon tissue can be evaluated in heath and disease. Furthermore, the difference between exercise deficiency due to an inadequate vascular bed, on the one hand, or a metabolic/genetic deficiency, on the other, is readily distinguished. These safe, economical and efficient evaluations of muscle performance are not ready for general and specialty medical evaluations without the use of large and confining magnets for NMR or radioactive materials for PET.